Sarcoidosis is a multisystem granulomatous disorder of unknown etiology that involves the lungs in over 90 percent of affected individuals and may cause end-stage fibrosis, cor pulmonale, and death. The pathologic hallmark of sarcoidosis is non-caseating granulomatous inflammation. Since extracts of diseased tissue injected intradermally elicit a nidus of granulomatous inflammation in patients with sarcoidosis that is indistinguishable from spontaneously arising granulomas (the Kveim reaction), we postulate that sarcoid tissue extracts contain disease-relevant antigens. Biophysical properties of the active component in Kveim extracts include relative heat stability, resistance to neutral detergents and proteases, and a dependence on tertiary structure. The overall goal of this application is to identify these pathogenic tissue antigens in sarcoidosis. Our central hypothesis is that sarcoidosis is caused by linked T and B cell immune responses to aggregates of altered proteins of microbial origin. Consistent with this hypothesis, our preliminary studies demonstrate the presence of a small number of protease-resistant, neutral-detergent insoluble proteins that by immunoblot analysis are targets of T cell dependent IgG from patients with sarcoidosis but not healthy controls. MALDI-TOF mass spectrometry and immunoblot analysis has identified the mycobacterial catalase-peroxidase protein from Mycobacterium tuberculosis (mKatG) or M. smegmatis in these protein fractions from sarcoidosis but not control tissues. Preliminary studies demonstrate both T and B cell responses to mKatG proteins in sarcoidosis, suggesting the mKatG proteins are relevant, pathogenic antigens in sarcoidosis. To test the hypothesis that mycobacterial KatG proteins are pathogenic antigens in sarcoidosis, we propose studies to determine the presence of mycobacterial KatG proteins in sarcoidosis and control tissues using MALDI-TOF mass spectrometry and protein immunoblot analyses. To determine whether these microbial proteins induce disease-specific immune responses, we will determine the molecular basis of the B and T cell immune responses to both M. tuberculosis and M. smegmatis KatG proteins and selected peptides, and determine whether mKatG proteins preferentially expand specific Valpha/Vbeta expressing T cells in patients with sarcoidosis and control subjects. Together, these studies offer the potential of identifying a specific group of microbial antigens involved in the pathogenesis of granulomatous inflammation in sarcoidosis, thus providing a novel target for therapy of this disease.